Abstract. Episialin (MUC1) is a transmembrane molecule with a large mucin-like extracellular domain protruding high above the cell surface. The molecule is located at the apical side of most glandular epithelial cells, whereas in carcinoma cells it is often present at the entire surface and it is frequently expressed in abnormally large quantities. We have previously shown that overexpression of episialin reduces cell-cell interactions. Here we show that the integrinmediated adhesion to extracellular matrix of transfectants of a melanoma cell line (A375), a transformed epithelial cell line (MDCK-ras-e) and a human breast epithelial cell line (HBL-100) is reduced by high levels of episialin. This reduction can be reversed by inducing high avidity of the ffl integrins by mAb TS2/16 (at least for/~l-mediated adhesion). The adhesion can also be restored by redistribution of episialin on the cell surface by monoclonal antibodies into patches or caps. Similarly, capping of episialin on ZR-75-1 breast carcinoma cells, growing in suspension, caused adherence and spreading of these cells.We propose that there is a delicate balance between adhesion and anti-adhesion forces in episialin expressing cells, which can be shifted towards adhesion by strengthening the integrin-mediated adhesion, or towards anti-adhesion by increasing the level of expression of episialin.

Episialin (MUC1, PEM, EMA, CA15-3 antigen) is a sialylated, membrane-associated glycoprotein with an extended mucin-like ectodomain. This domain mainly consists of 30-90 homologous 20-amino acid repeats that are rich in 0-glycosylation sites (serines and threonines). It is likely that this part forms a polyproline ,B-turn helix. As a result, the ectodomain can protrude more than 200 nm above the cell surface, whereas most cell surface molecules do not exceed a length of 35 nm. Normally, episialin is present at the apical side of glandular epithelial cells. On carcinoma cells, however, it can be strongly overexpressed and it is often present over the entire cell surface. We have previously shown that episialin, if it is interspersed between adhesion molecules, nonspecifically reduces cell-cell and cell-extracellular matrix interactions in vitro and in vivo, presumably by steric hindrance caused by the extreme length and high density of the episialin molecules at the cell surface. To analyze the molecular mechanism for this anti-adhesion effect in more detail, we have now deleted an increasing number of repeats in the episialin cDNA and transfected the resulting mutants into murine L929 cells expressing the homophilic adhesion molecule E-cadherin. Here we show that the length of episialin is the dominant factor that determines the inhibition of E-cadherin-mediated cell-cell interactions. For the anti-adhesive effect mediated by the full length episialin, charge repulsion by negatively charged sialylated 0-linked glycans is far less important.

The mucin-like glycoprotein episialin (MUC1) is highly overproduced by a number of human carcinomas. We have shown previously in a variety of mammalian cell lines that overexpression of this very large transmembrane molecule diminishes cellular adhesion, suggesting that episialin/MUC1 overexpression may play an important role in tumor invasion and metastasis. By using in situ hybridization, we show here that episialin/ MUC1 mRNA expression can be increased more than 10-fold in breast carcinoma cells relative to the expression in adjacent normal breast epithelium. In search of the molecular mechanism of this overexpression, we observed that the episialin/MUC1 promoter contains a candidate binding site for transcription factors of the STAT family ϳ500 base pairs upstream of the transcription start site. Cytokines and/or growth factors such as interleukin-6 or interferon-␥ can activate STATs. In the human breast carcinoma cell line T47D, both compounds are able to stimulate transcription of a luciferase reporter gene under the control of a 750-base pair MUC1 promoter fragment proximal to the transcription start site. The observed increase is entirely mediated by the single STAT-binding site, since mutation of this site abolishes stimulation of the reporter by interleukin-6 and interferon-␥. In addition, mutation of the STAT site also decreased the promoter activity in nonstimulated T47D cells, suggesting that the STAT-binding site is among the elements that are involved in the overexpression of MUC1 in tumor cells. Episialin/MUC11 (also known as MUC1, PEM, CA 15-3 antigen, or EMA) is a transmembrane molecule with a large extracellular mucin-like domain. In normal cells episialin/ MUC1 is exclusively present at the apical side of the cell, but in carcinoma cells normal polarization is lost and episialin/MUC1 co-localizes with adhesion molecules such as integrins and cadherins. The long and relatively rigid extracellular domain of episialin/MUC1 can shield these adhesion molecules and diminish cellular adhesion, if present at a sufficiently high density on the cell surface (1-3). Overexpression of episialin/MUC1 in carcinoma cells has been frequently reported (4 -6) and is expected to have a similar effect on cellular behavior as loss of E-cadherin, the major epithelial cell-cell adhesion molecule, which has been shown to promote invasion and metastasis of carcinoma cells (for review see Refs. 7-9). Therefore, we have proposed that episialin/MUC1 also plays an important role in invasion and metastasis in vivo (10). Indeed, transgenic mice overexpressing episialin/MUC1 develop more aggressive lung tumors than nontransgenic mice, 2 whereas episialin/MUC1 null mice show a slower rate of tumor progression (11).Recent reports have shown that episialin overexpression in various types of neoplasia correlates with poor survival (12)(13)(14). Episialin/MUC1 is also the antigen that is measured in the CA 15-3 assay (the main blood marker to detect recurrence of breast cancer), and it is a molecule that is widely considered as ...

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